The sequential, step-wise processing of slide surface mounted biological material is known and used in various procedures. Some of such procedures have come into widespread common usage for analytical and diagnostic purposes in various biological disciplines including biotechnology, biochemistry, molecular biology, molecular genetics, cytogenetics, cell biology, pharmacology, immunology, and the like. Such procedures are commonly carried out with each one of a series of processing steps involving the contacting of such mounted material with a variety of fluids (liquid and/or gases). Unless the particular processing step sequence is properly carried out, the results desired are not obtained. The total number of such processing steps used in any given procedural embodiment can vary, depending upon many variables.
Examples of such procedures include staining of tissue, cell, and cell-derived material, such as immunohistochemical staining of slide mounted biological tissue, and the like; hybridization of slide-bound nucleic acids with labeled probes that incorporate complementary nucleotide sequences, such as in situ hybridization; and the like.
In, for example, the staining of slide mounted specimens of biological materials, the prepared slides may be dipped successively into individual ones of a series of small vessels or jars, each about 0.2 to about 2 liters in volumetric capacity, and each containing a particular liquid treating composition, such as washing agents, buffers, denaturation agents, dehydrating agents, dyes and the like. One or more dye materials may be used to highlight different cells, structures of cells, intracellular structures, cellular products or the like. Resulting dyed slides may be washed and dried, possibly stored, and then microscopically examined. Selective staining procedures using specific antibody or gene probes have been developed and are advantageously highly specific and sensitive. However, such procedures require many successive steps to be carried out.
In, for another example, in situ hybridization, the presence, location, identity, or quantity of a particular nucleic acid type in a given cell structure is determined. Starting cytological preparations mounted on glass slides may be variously treated. For instance, one step may involve a treatment to denature the DNA or RNA (without losing the identity of the cellular nucleic acid structures). Thereafter, the resulting slide may be contacted with a liquid composition containing an oligonucleotide probe which is labeled with an appropriate radioisotope, fluorophore, enzyme, hapten, avidin, or the like. Hybridization occurs between the probe and the complementary nucleic acid, such as a gene on a chromosome, a ribosomal RNA (rRNA), or the like. The resulting slide is suitably washed to remove residual probes Subsequent other further procedural processing steps may also be undertaken. The resulting slide is examined using an evaluation procedure such as, for example, microscopy, autoradiography, fluorescence measurement, photon emission, or the like. Up to about 30 or more controlled step sequences may be involved for a single hybridization procedure.
Since multi-step processing of slide mounted biological materials typically involves a multiplicity of slides, a plurality of process liquids, and a plurality of step conditions, control problems arise in achieving identical treatment of all slides of a slide plurality which are all undergoing the same processing sequence An added complication is that some processing liquids are very costly and must be used sparingly, such as liquids containing nucleotide probes. Such liquids are employed under conditions which differ from the conditions employed in other steps in the processing sequence Such make sequential problems multi-step processing techniques difficult to carry out identically for a plurality of slides.
There is a need in the art for apparatus and methods which can be operated and practiced so as to carry out a given multi-step slide processing sequence in a replicable manner. All variables affecting a slide surface during each individual step of the sequence need to be controlled, including temperature, time, and the fluid contacting variables, such as reagent concentration, fluid flow rate, slide surface residence time, and the like. Typically, it is desired that all slides of a group thereof should receive substantially identical processing whether or not individual slides of such group are processed parallel to one another in a given procedure or are in series relationship to one another in successive procedures.
Further, there is a need in the art for apparatus and methods which permit such a controlled step sequence to be automatically carried out under preliminarily programmed conditions. Still further, there is a need in the art for such apparatus and methods which can be practiced, if desired, with minimum amounts of processing fluids and agents particularly with regard to probes.
So far as is now known, apparatus and methods with such capability have not previously been known to the art.